A seed sequence variant in miR-145-5p causes multisystem smooth muscle dysfunction syndrome
Authors: Christian Lacks Lino Cardenas, Lauren C Briere, David A Sweetser, Mark E Lindsay, Patricia L Musolino
Journal: Journal of Clinical Investigation (2023)

Multisystemic smooth muscle dysfunction syndrome (MSMDS, OMIM #613834) is an ultrarare smooth muscle myopathy (1).Cases are monogenic from missense variation at arginine 179 of the ACTA2 gene (1, 2).Herein, we describe a case of MSMDS associated with a single-nucleotide variant in the gene MIR145.Fetal ultrasound revealed polyhydramnios, enlarged abdomen and bladder, and prune belly syndrome.Profound gastrointestinal dysmotility was identified during infancy.His cerebrovascular disease began with frontal cortex and watershed strokes at approximately 2.5 years of age.Straightening of cerebral arteries and flattening of the genu of the corpus callosum and pons was observed.During school age he had multiple strokes consistent with arterial ischemic and watershed infarctions.Severe progressive steno-occlusive disease developed, which was worse in the anterior circulation (Figure 1, A andB).The vascular anatomy also showed straightening and decreased caliber of the terminal internal carotid artery, consistent with described cases of MSMDS (3, 4).Thoracic aortic imaging has been normal.A thoracic aortic aneurysm/dissection panel was negative, including analysis of ACTA2.Quad genome sequencing was negative; however, research-based analysis revealed a de novo single-nucleotide variant in MIR145 (NR_029686.1:n.18C>A) (Figure 1C).This variant is absent from gnomAD, has a CADD score of 20.9, and MIR145 is enriched in tissues with high smooth muscle cell (SMC) content (5) (Supplemental Figure 1; supplemental material available online with this article; https://doi.org/10.1172/JCI166497DS1).The MIR145 transcript is processed into 2 microRNAs (miRs), with the variant position at nucleotide 3 of miR-145-5p.To determine whether the miR-145-5p variant could mediate the observed patient phenotype of smooth muscle dysfunction, we undertook molecular analysis.Cases of MSMDS to date have been caused by recurrent missense variants in the ACTA2 gene, altering arginine 179 (2).These variants impair α-smooth muscle actin (α-SMA) function, resulting in a cellular state resembling a loss of protein function.The miR-145-5p variant is located within the seed sequence (nt 2-8), the portion of a miR that stalls lateral diffusion of the RISC complex and promotes stable interactions with complementary RNAs (Figure 1C).We hypothesized that mutant miR-145-5p may not be able to target 3′ UTRs that mediate proper SMC function and may thus result in cellular changes similar to ACTA2 R179 variants.To assess this possibility, we exposed human vascular SMCs to an siRNA targeting miR-145-5p, wild-type (WT) miR-145-5p, or a mutant version of miR-145-5p with the patient variant.Indeed, transfection of either an siRNA against miR-145-5p or mutant miR-145-5p induced a notable decrease in the expression of several cytoskeletal proteins, including transgelin, calponin, and importantly, α-SMA (Figure 1D and Supplemental Figure 2; see complete unedited blots in the supplemental material).Cellular models of the ACTA2 R179H mutation demonstrate global filamentous actin (F-actin) cytoskeletal deficien-